Skeletal muscle mitochondrial interactome remodeling is linked to functional decline in aged female mice.

Genomic, transcriptomic and proteomic approaches have been used to gain insight into molecular underpinnings of aging in laboratory animals and in humans. However, protein function in biological systems is under complex regulation and includes factors besides abundance levels, such as modifications, localization, conformation and protein-protein interactions. By making use of quantitative chemical cross-linking technologies, we show that changes in the muscle mitochondrial interactome contribute to mitochondrial functional decline in aging in female mice. Specifically, we identify age-related changes in protein cross-links relating to assembly of electron transport system complexes I and IV, activity of glutamate dehydrogenase, and coenzyme-A binding in fatty acid ß-oxidation and tricarboxylic acid cycle enzymes. These changes show a remarkable correlation with complex I respiration differences within the same young-old animal pairs. Each observed cross-link can serve as a protein conformational or protein-protein interaction probe in future studies, which will provide further molecular insights into commonly observed age-related phenotypic differences. Therefore, this data set could become a valuable resource for additional in-depth molecular studies that are needed to better understand complex age-related molecular changes.

From mitochondria to cells to humans: Targeting bioenergetics in aging and disease.

In vivo control over metabolism is at the cutting edge of biomedical research. The particulars of mitochondrial function are especially important to understand in vivo to progress metabolic therapies that will be relevant for diseases of aging. Understanding the differences between how mitochondria function in vitro versus in vivo will be a necessary challenge to overcome to achieve mitochondrial medicine. In this article we outline how discoveries in invertebrate models will be informative for understanding the basic biology of mitochondria to streamline translation to mammals and eventually to humans. Further, we highlight examples of how what is known about mitochondria in vitro is translatable to in vivo models and, in some cases, to human diseases.

Short-term Calorie Restriction and 17α-Estradiol Administration Elicit Divergent Effects on Proteostatic Processes and Protein Content in Metabolically Active Tissues.

17α-Estradiol (17α-E2) is a "non-feminizing" estrogen that extends life span in male, but not female, mice. We recently reported that 17α-E2 had robust beneficial effects on metabolic and inflammatory parameters in aged male mice. However, it remains unclear if 17α-E2 also delays other "hallmarks" of aging, particularly maintaining proteostasis. Here, we used isotope labeling methods in older mice to examine proteostatic mechanisms. We compared weight-matched mild calorie restricted (CR) and 17α-E2 treated male mice with the hypothesis that 17α-E2 would increase protein synthesis for somatic maintenance. 17α-E2 had no effect on protein synthesis or DNA synthesis in multiple tissues, including white adipose tissue. Conversely, mild short-term CR decreased DNA synthesis and increased the protein to DNA synthesis ratio in multiple tissues. Examination of individual protein synthesis and content did not differentiate treatments, although it provided insight into the regulation of protein content between tissues. Contrary to our hypothesis, we did not see the predicted differences in protein to DNA synthesis following 17α-E2 treatment. However, mild short-term CR elicited differences consistent with both lifelong CR and other treatments that curtail aging processes. These data indicated that despite similar maintenance of body mass, 17α-E2 and CR treatments elicit distinctly different proteostatic outcomes.

Insulin-like growth factor receptor signaling regulates working memory, mitochondrial metabolism, and amyloid-ß uptake in astrocytes.

OBJECTIVE: A decline in mitochondrial function and biogenesis as well as increased reactive oxygen species (ROS) are important determinants of aging. With advancing age, there is a concomitant reduction in circulating levels of insulin-like growth factor-1 (IGF-1) that is closely associated with neuronal aging and neurodegeneration. In this study, we investigated the effect of the decline in IGF-1 signaling with age on astrocyte mitochondrial metabolism and astrocyte function and its association with learning and memory. METHODS: Learning and memory was assessed using the radial arm water maze in young and old mice as well as tamoxifen-inducible astrocyte-specific knockout of IGFR (GFAP-CreTAM/igfrf/f). The impact of IGF-1 signaling on mitochondrial function was evaluated using primary astrocyte cultures from igfrf/f mice using AAV-Cre mediated knockdown using Oroboros respirometry and Seahorse assays. RESULTS: Our results indicate that a reduction in IGF-1 receptor (IGFR) expression with age is associated with decline in hippocampal-dependent learning and increased gliosis. Astrocyte-specific knockout of IGFR also induced impairments in working memory. Using primary astrocyte cultures, we show that reducing IGF-1 signaling via a 30-50% reduction IGFR expression, comparable to the physiological changes in IGF-1 that occur with age, significantly impaired ATP synthesis. IGFR deficient astrocytes also displayed altered mitochondrial structure and function and increased mitochondrial ROS production associated with the induction of an antioxidant response. However, IGFR deficient astrocytes were more sensitive to H2O2-induced cytotoxicity. Moreover, IGFR deficient astrocytes also showed significantly impaired glucose and Aß uptake, both critical functions of astrocytes in the brain. CONCLUSIONS: Regulation of astrocytic mitochondrial function and redox status by IGF-1 is essential to maintain astrocytic function and coordinate hippocampal-dependent spatial learning. Age-related astrocytic dysfunction caused by diminished IGF-1 signaling may contribute to the pathogenesis of Alzheimer's disease and other age-associated cognitive pathologies.

AG311, a small molecule inhibitor of complex I and hypoxia-induced HIF-1α stabilization.

Cancer cells have a unique metabolic profile and mitochondria have been shown to play an important role in chemoresistance, tumor progression and metastases. This unique profile can be exploited by mitochondrial-targeted anticancer therapies. A small anticancer molecule, AG311, was previously shown to possess anticancer and antimetastatic activity in two cancer mouse models and to induce mitochondrial depolarization. This study defines the molecular effects of AG311 on the mitochondria to elucidate its observed efficacy. AG311 was found to competitively inhibit complex I activity at the ubiquinone-binding site. Complex I as a target for AG311 was further established by measuring oxygen consumption rate in tumor tissue isolated from AG311-treated mice. Cotreatment of cells and animals with AG311 and dichloroacetate, a pyruvate dehydrogenase kinase inhibitor that increases oxidative metabolism, resulted in synergistic cell kill and reduced tumor growth. The inhibition of mitochondrial oxygen consumption by AG311 was found to reduce HIF-1α stabilization by increasing oxygen tension in hypoxic conditions. Taken together, these results suggest that AG311 at least partially mediates its antitumor effect through inhibition of complex I, which could be exploited in its use as an anticancer agent.